Signal Transducer and Activator of Transcription (STAT) proteins are a family of transcriptional activators essential for many growth factor signalled responses. Stat4 is specifically activated by interleukin (IL)-12 and has been implicated in biological responses including proliferation of activated T cells, interferon (IFN)gamma production from NK and T cells, increased cytotoxicity in NK and T cells and development of T helper (Th) type l effector cells. It is not well understood how Stat4 regulates these processes. Using Stat4-deficient mice as a model, this proposal will elucidate the mechanisms by which Stat4 functions. IL-12 does not induce lymphocyte proliferation in the absence of Stat4, a defect correlated with dysregulation of a cyclin dependent kinase inhibitor, p27(Kip1). We will determine how p 27(Kip1)is regulated by Stat4 and determine if p27(Kip1) dysregulation is the only defect using Stat4-p27(Kip1) double deficient mice. To determine how Stat4 functions as a transcriptional activator, we will examine IL-12 induced transcriptional events-using the cD25 gene as a model in wildtype and Stat4-deficient T cells. We will further explore the domains of Stat4 involved in transcriptional activation by generating mice transgenic for mutant Stat4 proteins and analyze in vivo function of the mutant proteins in Stat4-expressing and - deficient cells. Finally, we will examine the role of Stat4 in programming cells to the Th1 phenotype by studying granuloma formation and hematopoiesis, two in vivo systems of Stat4/Th1 function, using mice that are wildtype, Stat4-deficient (which lack Th1 cells) and Stat6-deficient (which have only Th1 cells and lack Th2 cells). These studies will lead to a better understanding of Stat4 function in vivo. They will elucidate how STAT proteins regulate gene transcription, differentiation and contribute to lymphocyte proliferation, which will be important in determining the role of STATs in leukemia and lymphoma. They will also demonstrate how Stat4 is important in programming Th1 activity in vivo and how Th1 regulated processes such as inflammation and hematopoiesis are controlled. A better understanding of Th1 function will provide biological targets for treatment of chronic inflammation and autoimmunity.